1.7.3.3: factor-independent urate hydroxylase

This is an abbreviated version!
For detailed information about factor-independent urate hydroxylase, go to the full flat file.

Word Map on EC 1.7.3.3

Reaction

Urate
+
O2
 +
H2O
=
5-hydroxyisourate
 +
H2O2

Synonyms

AaUO, AgUOX, ELITEK, Fasturtec, N-35, Nodule specific uricase, Nodulin 35, Nodulin 35 homolog, Non-symbiotic uricase, oxidase, urate, Rasburicase, Uaz, Uox, Urate oxidase, urate oxidoreductase, UriA, uric acid oxidase, uricase, uricase II, Uricoenzyme, Uricozyme

ECTree

     1 Oxidoreductases
         1.7 Acting on other nitrogenous compounds as donors
             1.7.3 With oxygen as acceptor
                1.7.3.3 factor-independent urate hydroxylase

Engineering

Engineering on EC 1.7.3.3 - factor-independent urate hydroxylase

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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D307R
-
mutant is an inactive homotetramer of lower solubility
E318R
-
mutant is similar to S314Stop
L171I/Y182F/Y187F/A193S
-
mutant displays higher activity and lower thermostability
L322D
-
mutant behaves similar to wild-type
L322R
-
mutant shows reduced activity and thermal stability at pH 7.4, enhanced stability at pH 9.2
R310E
-
displays the lowest stability at pH 7.4 or 9.2 among tested mutants and the strongest effects of pH values on thermal stability
S314Stop
-
mutant is a homotetramer of about 25% activity, 5 % half-life at pH 7.4 but 100 % half-life at pH 9.2
V144A
-
mutant displays higher activity and consistent thermostability
Y249F
-
thermal stability is reduced by 50fold at pH 7.4, by about 7fold at pH 9.2
Y319F
-
thermal stability is reduced by one magnitude at pH 7.4, but by 2fold at pH 9.2
Y319F/Y249F
-
mutant is an inactive homotetramer
D307R
-
mutant is an inactive homotetramer of lower solubility
-
R310E
-
displays the lowest stability at pH 7.4 or 9.2 among tested mutants and the strongest effects of pH values on thermal stability
-
Y249F
-
thermal stability is reduced by 50fold at pH 7.4, by about 7fold at pH 9.2
-
Y319F
-
thermal stability is reduced by one magnitude at pH 7.4, but by 2fold at pH 9.2
-
D79A
-
the KM-value for urate is 82% of the wild-type value
F179A
-
decreases Vmax by 2 orders of magnitude
F179Y
-
mutation decreases Vmax by 2-fold
T69A
-
mutant enzyme has a maximal velocity of 3% of the wild-type value. Ionization at pH 6.4 that is observed with the wild-type enzyme is absent in the mutant. The KM-value for urate is 5fold higher than that of the wild-type enzyme
T69A/K9M
-
the KM-value for urate is 16.1fold higher than that of the wild-type enzyme
T69V
-
the KM-value for urate is 30.9fold higher than that of the wild-type enzyme
A296V
about 50% ioncrease in catalytic efficiency
A89T/G91A7V92M/H245L/E252A/M253I/R291K/A296V/A301S/K303R
replacement with the corresponding residues of human enzyme. About 30% of wild-type catalytic efficiency
H245L/E252A/M253I/R291K/A296V/A301S/K303R
replacement with the corresponding residues of human enzyme. Catalytic efficiency is higher than in wild-type, but below the efficiency of mutant R291K/A296V/A301S/K303R
I115V/H119R/L120F/H245L/E252A/M253I/R291K/A296V/A301S/K303R
replacement with the corresponding residues of human enzyme, complete loss of activity
R119H
complete loss of activity
R291K/A296V/A301S/K303R
replacement with the corresponding residues of human enzyme. About 50% increase in catalytic efficiency
synthesis
-
expression of baboon uricase gene attached with Trx and hexahistidine tags in Escherichia coli Rosetta (DE3). The final yield of mature baboon uricase is 136.0 mg/l with enzyme activity of 17.93 U/mg
additional information